Hematopoietic Stem Cells (HSCs) are the only cells responsible for sustained endogenous hematopoiesis, and early and sustained bone marrow transplant engraftment. While HSCs under many conditions can self-renew and expand their numbers dramatically in vivo, this has not been recapitulated in vitro. The objective of this PPG is to use a multidisciplinary approach to identify genes that might regulate stem cell fates (such as self-renewal vs. differentiation). Four collaborating laboratories have brought complementing systems and technologies for this purpose. The I. Weissman lab has developed sensitive assays to identify genes expressed in HSCs, and to assay candidate genes transfected to ES cells by following their HSC progeny in vivo. M. Clarke has identified a genetic polymorphism that results in high vs. Low levels of HSCs in two mouse strains, and will attempt positional cloning of a candidate gene. L. Hood will obtain full complexity cDNA libraries (from M. Clarke) that represent each HSC subset, and will use genomics approached and high density cDNA arrays to identify candidates for further study. M. Fuller has identified 3 Drosophila genes expressed in hematopoietic and/or male germline progenitors that regulated spermatogonial stem cell fates, and has evidence they may be conserved in rodents. By coordinating gene candidate searches from all 4 labs with the Hood lab high density arrays and informatics capabilities, high priority candidate genes will be chosen to be tested biologically in the Weissman lab. The long-term objective of finding a HSC self-renewal pathway should enable future protocols for HSC expansion in vivo or in vitro, which will be valuable in humans for autografting, for gene therapy in cycling HSCs, and for allografting HSCs (without T cells) for the induction of tolerance for organ transplants, and for replacement of autoimmune hematolymphoid systems.